《植物生理学报》 2014, 50(8): 1184-1194

胡萝卜AP2/ERF-B1亚族两个转录因子基因的克隆及其非生物胁迫响应分析

黄蔚, 王枫, 谭国飞, 徐志胜, 李梦瑶, 熊爱生*

作物遗传与种质创新国家重点实验室, 农业部华东地区园艺作物生物学与种质创制重点实验室, 南京农业大学园艺学院, 南京210095

通信作者：熊爱生；E-mail: xiongaisheng@njau.edu.cn；Tel: 025-84396790

摘　要：

AP2/ERF是植物中普遍存在的一类重要转录因子, 参与植物整个生命周期的生长发育和逆境信号转导。本研究以胡萝卜(Daucus carota) ‘黑田五寸’为试验材料, 基于其转录组和基因组数据, 检索和拼接获得胡萝卜AP2/ERF家族2个转录因子基因序列g39811和g47170。采用RT-PCR方法, 分别从‘黑田五寸’中克隆DcERF-B1-1 (g39811)和DcERF-B1-2 (g47170)转录因子基因。序列分析显示, 胡萝卜DcERF-B1-1和DcERF-B1-2转录因子基因分别含有630个和594个开放阅读框, 分别编码209和197个氨基酸; 均含有相对保守的AP2结合域, 具有典型的植物AP2/ERF类转录因子特征。从氨基酸组成成分、理化性质、亲水性/疏水性和三级结构上分析显示, 胡萝卜DcERF-B1-1和DcERF-B1-2转录因子亲水性大于疏水性, 其氨基酸序列可能属于亲水性蛋白。空间结构分析显示, 它们都具有1个α螺旋和3个β折叠。进化树分析显示, 二者均属于AP2/ERF家族转录因子中ERF亚族的B1组。实时定量荧光PCR显示, 在低温、干旱、盐胁迫的条件下, DcERF-B1-2转录因子比DcERF-B1-1转录因子对逆境的响应更大; 在高温的条件下, DcERF-B1-1转录因子比DcERF-B1-2转录因子对逆境的响应更大。

关键词：AP2/ERF; 胡萝卜; 转录因子; ERF亚族; 表达调控

收稿：2014-04-14   修定：2014-06-28

资助：教育部新世纪优秀人才支持计划(NCET-11-0670)、国家自然科学基金项目(31272175)、江苏省杰出青年基金项目(BK20130027)、中国博士后科学基金项目(2014M551609)、江苏高校优势学科建设项(2011PAPD)和江苏省双创计划(2011JSSC)。

Isolation and Analysis of Expression Profiles under Abiotic Stress Treatments of Two AP2/ERF-B1 Group Transcription Factor Genes from Daucus carota

HUANG Wei, WANG Feng, TAN Guo-Fei, XU Zhi-Sheng, LI Meng-Yao, XIONG Ai-Sheng*

College of Horticulture, Nanjing Agricultural University, State Key Laboratory of Crop Genetics and Germplasm Enhancement, Ministry of Agriculture Key Laboratory of Biology and Germplasm Enhancement of Horticultural Crops in East China, Nanjing, 210095, China

Corresponding author: XIONG Ai-Sheng; E-mail: xiongaisheng@njau.edu.cn; Tel: 025-84396790

Abstract:

AP2/ERF is an important family of transcription factors in higher plant, which encoded proteins involved in the development and signal transduction of abiotic/biotic resistance in higher plant. The g39811 and g47170 gene sequences, which encoding two AP2/ERF transcription factors, were isolated based on transcriptome and genome sequence data of carrot (Daucus carota). The DcERF-B1-1 (g39811) and DcERF-B1-2 (g47170) genes were cloned by RT-PCR method using cDNA as template from carrot cultivar ‘Heitianwucun’. Then, nucleic acid and deduced amino acid sequence, phylogenetic tree, and molecular modeling were predicted and analyzed. Sequence analysis indicated that the length of DcERF-B1-1 and DcERF-B1-2 genes were 630 bp and 594 bp, which encoded 209 and 197 amino acids, respectively. The DcERF-B1-1 and DcERF-B1-2 factors contained AP2-domain, which mainly found in higher plants AP2/ERF family factors. The DcERF-B1-1 and DcERF-B1-2 transcription factors maybe belong to hydrophilic protein. Protein three-dimensional structure prediction and analysis showed that the AP2-domain of DcERF-B1-1 and DcERF-B1-2 had one α-helix and three β-sheets. The DcERF-B1-1 and DcERF-B1-2 factors from carrot were classified into B1 subgroup of ERF subfamily transcription factor, which belongs to AP2/ERF family. Quantitative real-time PCR analysis of the expression profiles in carrot showed that the DcERF-B1-2 responded more severely than the DcERF-B1-1 by low temperature, high-salinity and salt stress treatments, and the DcERF-B1-1 responded more severely than the DcERF-B1-2 by high temperature treatment.

Key words: AP2/ERF; Daucus carota; transcription factor; ERF subfamily; expression profile